Amplification and Cloning of NS3 Gene of Dengue Virus

The RNA of dengue 2 virus (New Guinea C strain, NGC) was extracted from small amount of dengue 2 virus culture supernatant. The large NS3 gene fragment was amplified from RNA of dengue 2 virus with RT PCR method. The amplified NS3 gene fragment was cloned directly into the pUC18 plasmid vector. The recombinant plasmid was identified by agarose gel electrophoresis analysis after digestion with restriction endonuclease and nested PCR method. The results showed that the amplified NS3 gene fragment was about 1978 bp in length; the recombinant plasmid contained NS3 gene of dengue 2 virus. The method of amplified large fragment of dengue virus from small amount of dengue virus culture supernatant was established.